基因芯片用于组分中药新双龙方的配伍机制研究

采用基因芯片技术研究了中药新双龙方及其有效组分人参总皂苷和丹参总酚酸治疗急性心肌梗塞(AMI)大鼠的作用机制. 以结扎大鼠冠脉左前降支的方法制备急性心梗模型, 采用大鼠全基因组表达谱芯片分别筛选出AMI大鼠给药前及新双龙方与组分给药后的差异表达基因(Ratio>2或<0.5), 通过对4组差异基因的综合分析, 比较了新双龙方与组分作用方式的异同, 从分子调控水平上探讨复方配伍后的增效作用, 同时对与药效作用机理直接相关的钙离子转运相关的Cacna1d基因和血管修复相关的Flnb基因进行了实时荧光逆转录聚合酶链式反应(RT-PCR)定量. 结果表明, 新双龙方与2种有效组分对心梗大鼠基因表达的影响不同. 对差异基因的聚类分析以及药物的调控作用比较结果均表明复方比组分的疗效更好; 通路分析结果表明, 新双龙方与组分均主要通过调控钙信号转导通路、 丝裂原活化蛋白激酶信号转导通路(MAPK)信号等通路发挥药效作用, 且复方涉及更多通路; 筛选出了复方与组分共同调控的靶点基因, 其在3组中具有不完全相同的表达趋势, 但复方具有更积极的调控作用. 本研究结果从分子调控角度证实了复方新双龙方的配伍优势.     

新型六方铜铝磷酸盐的合成及苯酚催化羟化反应活性的研究

本文采用静态水热合成法合成了一种非水溶性固体铜铝磷酸盐(CuAlPO).研究结果发现,CuAlPO是一种与六方二氧化硅(SiO₂)晶体结构相同的新型铜铝磷盐,在温和反应条件下(30℃),CuAlPO具有较高的苯酚催化羟化反应活性.     

基于Tet-On 3G的IFITM3诱导表达MDCK细胞系的建立及功能分析

利用Tet-On 3G系统构建了含人IFITM3基因的真核表达质粒pTRE3G-IFITM3,并将其与调控质粒pLVX-Tet3G共转染犬肾细胞(MDCK),转染后48 h用G418和嘌呤霉素进行筛选,用多西环素(Dox)对获得的细胞系进行诱导表达鉴定,并进行Dox敏感性分析、IFITM3~+细胞百分数及定位分析.用含萤光素酶(Luciferase)报告基因的禽流感病毒H5/H7蛋白或VSV G蛋白包裹的假型慢病毒颗粒进行感染实验,检测萤光素酶活性.结果表明,筛选获得了携带人IFITM3的MDCK诱导表达细胞系,且IFITM3表达量与Dox剂量和诱导时间相关;确定Dox工作浓度为2.5μg/mL,诱导12 h时IFITM3~+细胞数达75%以上,且IFITM3在晚期内体/溶酶体存在分布;假型病毒感染及萤光素酶活性分析表明,IFITM3可显著抑制禽流感病毒H5,H7和VSV G蛋白介导的病毒进入,为深入探究其具体的抑制机制奠定了基础.     

钝顶螺旋藻Spirulina platensis两种不同形态藻丝体的光合作用和蛋白质差异表达分析

从钝顶螺旋藻Spirulina platensis中分离纯化获得了螺旋形和直线形两种不同形态的藻丝体. 通过对二者光合作用的研究发现, 螺旋形藻丝体具有较高的光饱和光合作用速率(P_m^Chla)和光饱和点, 比直线形藻丝体更能适应较高光强的环境; 而直线形藻丝体具有较低的光补偿点, 能在更低的光强下进行光合作用. 通过双向凝胶电泳对两种不同形态藻丝体的总蛋白进行比较分析, 从中找出了9个差异表达的蛋白点. 应用基质辅助激光解吸电离质谱对差异表达的蛋白质点进行鉴定和数据库检索, 结果表明, 3个蛋白与肽聚糖代谢有关, 2个蛋白与光合作用有关, 1个蛋白与细胞分裂调控相关, 1个蛋白为外膜通道蛋白, 2个蛋白为功能未知的假想蛋白.     

Identification of key genes and expression profiles in osteoarthritis by co-expressed network analysis

BackgroundThe underlying molecular characteristics of osteoarthritis (OA), a common age-related joint disease, remains elusive. Here, we aimed to identify potential early diagnostic biomarkers and elucidate underlying mechanisms of OA using weighted gene co-expression network analysis (WGCNA).Material and methodsWe obtained the gene expression profile dataset GSE55235, GSE55457, and GSE55584, from the Gene Expression Omnibus. WGCNA was used to investigate the changes in co-expressed genes between normal and OA synovial membrane samples. Modules that were highly correlated to OA were subjected to functional enrichment analysis using the R clusterProfiler package. Differentially expressed genes (DEGs) between the two samples were screened using the “limma” package in R. A Venn diagram was constructed to intersect the genes in significant modules and DEGs. RT -PCR was used to further verify the hub gene expression levels between normal and OA samples.ResultsThe preserved significant module was found to be highly associated with OA development and progression (P < 1e-200, correlation = 0.92). Functional enrichment analysis suggested that the antiquewhite4 module was highly correlated to FoxO signaling pathway, and the metabolism of fatty acids and 2-oxocarboxylic acid. A total of 13 hub genes were identified based on significant module network topology and DEG analysis, and RT-PCR confirmed that these genes were significantly increased in OA samples compared with that in normal samples.ConclusionsWe identified 13 hub genes correlated to the development and progression of OA, which may provide new biomarkers and drug targets for OA.     

Bioinformatics analysis and verification of gene targets for renal clear cell carcinoma

BackgroundIt is estimated that there are 338,000 new renal-cell carcinoma releases every year in the world. Renal cell carcinoma (RCC) is a heterogeneous tumor, of which more than 70% is clear cell renal cell carcinoma (ccRCC). It is estimated that about 30% of new renal-cell carcinoma patients have metastases at the time of diagnosis. However, the pathogenesis of renal clear cell carcinoma has not been elucidated. Therefore, it is necessary to further study the pathogenesis of ccRCC.MethodsTwo expression profiling datasets (GSE68417, GSE71963) were downloaded from the GEO database. Differentially expressed genes (DEGs) between ccRCC and normal tissue samples were identified by GEO2R. Functional enrichment analysis was made by the DAVID tool. Protein-protein interaction (PPI) network was constructed. The hub genes were excavated. The clustering analysis of expression level of hub genes was performed by UCSC (University of California Santa Cruz) Xena database. The hub gene on overall survival rate (OS) in patients with ccRCC was performed by Kaplan-Meier Plotter. Finally, we used the ccRCC renal tissue samples to verify the hub genes.Results1182 common DEGs between the two datasets were identified. The results of GO and KEGG analysis revealed that variations in were predominantly enriched in intracellular signaling cascade, oxidation reduction, intrinsic to membrane, integral to membrane, nucleoside binding, purine nucleoside binding, pathways in cancer, focal adhesion, cell adhesion molecules. 10 hub genes ITGAX, CD86, LY86, TLR2, TYROBP, FCGR2A, FCGR2B, PTPRC, ITGB2, ITGAM were identified. FCGR2B and TYROBP were negatively correlated with the overall survival rate in patients with ccRCC (P < 0.05). RT-qPCR analysis showed that the relative expression levels of CD86, FCGR2A, FCGR2B, TYROBP, LY86, and TLR2 were significantly higher in ccRCC samples, compared with the adjacent renal tissue groups.ConclusionsIn summary, bioinformatics technology could be a useful tool to predict the progression of ccRCC. In addition, there are DEGs between ccRCC tumor tissue and normal renal tissue, and these DEGs might be considered as biomarkers for ccRCC.     

聚类分析法研究化妆品的真伪

本文以活性炭吸附丝采集“永芳Ｆ真珠膏”的挥发性组份,用ＧＣ－９Ａ的气相色谱仪进行测定,对２４个真品进行Ｒ型聚类分析和因子分析,提取了１４个特征指标,并以Ｑ型聚类方式建立了鉴别“永芳Ｆ真珠膏”真伪的数学模型,用３个假冒产品和２个真品对模型进行验证,结果令人满意。     

高效液相色谱-电喷雾离子阱质谱法鉴定复方丹参方化学及代谢成分

应用高效液相色谱-电喷雾离子阱质谱法鉴定了复方丹参方和其在家兔血浆、尿及粪便中的代谢产物。实验采用ZorbaxC18色谱柱,二元线性梯度洗脱进行色谱分离,并与电喷雾质谱联用,根据正离子和负离子模式下的分子离子峰获得化合物分子量信息,通过与文献数据或部分对照品对照,确定化合物的可能结构。     

鲨鱼软骨质量的聚类分析法

用电感耦合等离子体原子发射光谱法（ＩＣＰ－ＡＥＳ）测定了１８个鲨鱼软骨真品中１３个元素含量，对测定结果进行分析和变量聚类分析，抽提５个特征指标，并以谱系数聚类建立了鉴别鲨鱼软骨真伪的数学模式，用３个假冒产品和一个鲨鱼软骨胶囊对其进行检验，结果表明，用无机元素所建立的数学模式不仅能鉴别鲨鱼软骨的真伪，还能区分鲨鱼软骨的部分及不同部位的混合物。     

Exploring the mechanism of action Xianlingubao Prescription in the treatment of osteoporosis by network pharmacology

In this study, the network pharmacology analysis method was used to explore the bioactive components and targets of Xianlinggubao (XLGB) and further elucidate its potential biological mechanisms of action in the treatment of osteoporosis (OP). The bioactive compounds and predictive targets of XLGB were collected from the traditional Chinese medicine systems pharmacology databases and analysis platform(TCMSP), the Encyclopeida of traditional Chinese medicine (ETCM), traditional Chinese medicine Databse@Taiwan, ChEMBL, STITCH, and SymMap database. The targets corresponding to OP were obtained by using Online Mendelian Inheritance in Man® (OMIM), GeneCards, the National Center for Biotechnology Information-Gene database. The XLGB-OP targets were obtained by intersecting with the targets of XLGB and OP. Protien-Protien interaciton (PPI) network was constructed using STRING online database and analyzed using Cytoscape 3.7.0 software to screen out hub genes. Gene ontology (GO) and KEGG enrichment analysis of the target in the PPI network was conducted using the ClusterProfiler package in R with adjusted p-value<0.05. A total of 65 XLGB bioactive compounds were screened corresponding to 776 XLGB targets and 2556 OP targets. The GO analysis and KEGG enrichment analyses suggested XLGB played a therapeutic roles in OP treatment via the interleukin-17 signaling pathway, hypoxia-inducible factor-1 signaling pathway, insulin resistance, Th-17 signaling pathway, etc. Five hub genes (AKT1, MAPK1, MAPK8, TP53, and STAT3) were screened using the degree algorithm, and molecular docking stimulation results showed that most bioactive compounds of XLGB had strong binding efficiency with hub genes. Overall, this study laid the foundation for further in vivo and in vitro experimental research and expanded the clinical applications of XLGB.     

HPLC法测定茶叶水提液中五种儿茶素和咖啡碱及其用于茶叶分类的研究

建立高效液相色谱/二极管阵列检测器(HPLC/DAD)同时测定茶叶中(-)-没食子儿茶素(GC),(-)-表没食子儿茶素(EGC),(-)-表没食子儿茶素没食子酸酯(EGCG),(-)-表儿茶素(EC),(-)-表儿茶素没食子酸酯(ECG),咖啡碱(caffeine)6种组分的分析方法,并采用聚类分析探讨以这6种活性成分为指标对茶叶进行分类的方法。采用C18柱,甲醇和0.05%三氟乙酸水溶液为流动相,梯度洗脱,DAD双波长(210、278 nm)同时检测,采用标准物质保留时间和电喷雾飞行时间质谱(ESI TOF-MS)双重定性。结果表明,各组分的色谱峰均达到基线分离,在210 nm对(-)-没食子儿茶素(GC)定量,278 nm对其它组分定量准确。该法重复性好,灵敏度高,回收率高,已用于不同种类的33种实际茶叶样品的测定。以这6种活性成分的含量为指标,采用聚类分析法可对33个红茶、黑茶、绿茶、乌龙茶样本进行合理分类,并能反映茶叶品质的差异。